Transforming growth factor-β (TGF-β) acts as an important regulator of homeostasis in mature tissues by promoting growth inhibitory and cell death processes. Nevertheless, deregulated TGF-β activity results in severe disease pathologies. For example, during the course of cancer progression, cancerous cells frequently lose their responsiveness to TGF-β-mediated growth inhibition and TGF-β becomes a promoter of cancer, largely due to its enhancement of metastasis, immunosuppression and angiogenesis. In other cases, TGF-β overexpression leads to fibrotic disorders due to abnormal extracellular matrix accumulation. Indeed, the TGF-(3 superfamily consists of a large group of cytokines (e.g. activin, myostatin, BMPs, nodal) which when deregulated give rise to multiple disease states (Gordon 2008). A similar situation exists for particular ligands from other families, e.g. upregulated Sonic Hedgehog and Delta/Notch signaling are highly implicated in several different cancers, including gliomas (Li 2009). There is therefore a growing need for cytokine antagonists.
The most successful biologic therapeutics on the market function by antagonizing receptor-ligand interactions, e.g. antibodies that target and block the receptor or ligand. Receptor ectodomain-based ligand traps are a new class of therapeutics that, like antibodies, can bind and neutralize ligands, but have the advantage of being optimized more readily using protein engineering approaches. Dimerization of receptor ectodomains is of particular importance for promoting increased ligand trapping potency by providing a bridged-binding avidity effect. Dimerization can be achieved by fusing an ectodomain to the Fc portion of IgG. Several receptor Fc traps, including TGF-β RII-Fc and activin RII-Fc, are currently being evaluated in preclinical or clinical trials and four have been FDA-approved as therapeutic drugs (Huang 2009). A de novo designed heterodimerizing coiled-coil peptide system has been developed as an alternative dimerization approach to generate homobivalent and heterovalent TGF-β receptor traps that exhibit TGF-β neutralization IC50s in the low nM range (De Crescenzo 2004; De Crescenzo 2008). These coiled-coil traps have the advantage of being smaller than antibodies and Fc fused traps thus improving their tissue penetration.
Although widely used as therapeutics, monoclonal antibodies 1) are less amenable to optimization through protein engineering approaches since they are composed of heavy and light chains, 2) require complex manufacturing and 3) are large molecules (about 180 KDa) thus limiting tissue penetration. In contrast, receptor-Fc traps and coiled-coil receptor traps are more readily engineered and produced, and are smaller (about 120 KDa and about 80 KDa, respectively). Furthermore, in the case where a heterobivalent receptor trap is desired, assembly using the heterodimerizing E and K coil system has the advantage of potentially being able to promote 100% formation of heterodimers. This is accomplished by coexpressing two fusion proteins, for example receptor A-Ecoil and receptor B-Kcoil, in the producer cells. This is not the case when using the Fc homodimerization moiety where co-production of receptor A-Fc and receptor B-Fc theoretically results in receptor dimer combinations in the following proportions: 25% AA, 25% BB and 50% AB, which would subsequently require purification from each other. Nevertheless, two drawbacks of the coiled-coil system are 1) the non-covalent nature of coil dimerization may lead to separation of the receptor chains in the blood of the injected host, hence reducing trap potency and 2) the artificial coils may be immunogenic.
There is a need in the art for receptor-based traps that have one or more of the advantages of present receptor-based traps while minimizing one or more of the disadvantages.